1) Field of the Invention
The present invention relates to an apparatus and method for injecting a substance into cells.
2) Description of the Related Art
To modify the characteristics of the cells it is common to inject genes or drugs into cells in the field of life science, particularly regeneration medicine and genome drug discovery. With such a technology, roles of genes can be made clear and tailor made medicines that perform gene therapy suited for genetic characteristics of an individual can be prescribed.
Various methods have been proposed for injecting liquids into the cells to produce genetically modified cells. These methods include electrical methods, chemical methods, mechanical methods, and optical methods. The electropolation method is an example of the electrical method. The lipofection method is an example of the chemical method. The microinjection method is an example of the mechanical method. The laser injection method is an example of the optical method.
Among these, the electropolation method includes puncturing a cell membrane with electric pulses to form a hole in the cell membrane and injecting a drug fluid through the hole. This method can be used for any combination of cells and drug fluids. However, it is difficult to accurately control the puncturing of the cell. For example, the cell membrane may not be punctured successfully or, conversely, the cell membrane is punctured to the extent that the cells may die. There is also a problem that the ratio of successful introduction of the substance in the cell is very low.
The lipofection method involves mixing charged liposomes with DNA and allowing the resultant to adsorb onto the surface of cells so that the DNA enters into the cells. The virus vector method includes incorporating a gene into a virus DNA to prepare a genetic recombinant virus, and introducing the gene into the cells with the mechanism of the infection of the genetic recombinant virus.
However, the chemical and biological methods can be applied only to the limited combinations of the cells and the substances. In particular, in the case of the virus vector method, the risk of causing infectious diseases is relatively high since the method uses cells that have a high infectious capacity.
The microinjection method includes filling a drug fluid in a fine needle of diameter 1 micrometer (μm) and jabbing the needle into a cell and injecting the drug fluid in the cell (see, for example, Japanese Patent Application Laid-Open Publication No. H08-290377). FIG. 21 is a schematic for explaining a method for injecting a drug fluid into cells by the microinjection method. A needle 1 that holds a drug fluid 2 is jabbed into a cell membrane of a cell 3, and a pressure is applied to the drug fluid 2 so that the drug fluid 2 is injected into the cell 3 in the direction toward a nucleus 4 of the cell 3.
The tip of the needle 1 has a high surface tension because it is very thin, therefore, the drug fluid 2 is difficult to be led to the tip. One approach is to insert a glass rod 5 inside the needle 1. The presence of the glass rod 5 breaks the balance of the surface tension at an acute angle portion formed between the glass rod 5 and an inside of the needle 1, allowing the drug fluid 2 to be led to the tip of the needle 1.
The laser injection method involves dissolving a drug in a culture fluid for culturing cells and irradiating a laser beam to the cell membranes of the cells to form an opening in the cell membranes without using any needle. FIG. 22 is a schematic for explaining a method for injecting a drug fluid by the laser injection method. The drug fluid 2 is caused to penetrate into a cell 7 by the Brownian motion through an opening that is formed by the irradiation of a laser 6 to the cell 7 (see, for example, Japanese Patent Application Laid-Open Publication No. 2003-70468).
In the conventional microinjection method, if the same needle is used repeatedly, pieces of the cell membrane stick to the needle or enter into the needle. The pieces of the cell membrane that stick to the needle make the tip of the needle thicker. The pieces of the cell membrane that enter into the needle clog the tip. These situations make the injection difficult. Accordingly, it becomes necessary to periodically exchange the needle with a new one.
On the other hand, in the conventional laser injection method, it is necessary to make the laser beam very thin to precisely control the irradiation position of the laser so as not to damage the cells. For this purpose, it becomes necessary to observe the cells through a high magnification objective lens, which decreases the efficiency of the method.
The drug fluid is expensive and cannot be used in large amounts. This makes it necessary to use a smaller vessel to charge the cells to avoid a decrease in the concentration of the drug fluid. Accordingly, when the drug fluid is to be injected into a lot of cells, the operation of injecting the drug fluid into the cells and the operation of recovering the injected cells must be performed, for example, by charging the cells in separate vessels, respectively, one for one. This further decreases the efficiency of the conventional methods.